Receptors for the glycoprotein hormone follicle stimulating hormone (follitropin; FSH) are present in the Sertoli cells of testis and granulosa cells of ovaries. FSH is essential for normal high quality gametogenesis in males and females. Glycoprotein hormones which function in fertility and normal thyroid function are of general medical significance. Function of the FSH receptor (FSHR) beyond the initial binding event is regulated by cytoplasmic proteins which bind to receptor intracellular loops (il_) and regulate receptor function and act to organize receptors. Adapter proteins that have been identified which interact with human FSHR (hFSHR) are the focus of these studies. Aim one is to study the role of newly discovered adapter proteins APPL 1 and 2, that bind to FSHR-iLL Using siRNA to inhibit their synthesis in primary cultures of rat granulosa cells, the role of APPL1 and APPL2 regulation in FSH driven granulosa cell functions will be determined. In addition, amino acids in FSHR-IL1 which are essential for interaction with APPL proteins will be determined so that binding defective mutants can be studied for trafficking defects. Aim two will study the adapter protein 14-3- 3 tau which binds to FSHR-iL.2. It will be determined how inhibition of 14-3-3 tau affects FSHR signal transduction pathways in primary cultures of rat granulosa cells. Mutants ofhFSHR-il_2 defective in binding to 14-3-3 tau will be prepared and FSH activation of the Erk pathway will be studied in rat granulosa cell lines expressing these FSHR mutants. The third aim will determine the functional significance of the protein kinase CK2 (PKCK2) consensus site of thehFSHR-il_3. Consequences of inhibition of PKCK2 on steroidogenesis in primary cultures of human granulosa cells will be studied. It will be determined if PKCK2 interacts with hFSHR-il_3,and if consensus site mutants of the receptor are ubiquitination defective. Phosphorylation of hFSHR by PKCK2 will also be studied. The overall goal of the studies is to provide new information to better our understanding of regulation of gonadal function by gonadotropins and provide new avenues for fertility